5 research outputs found
Mechanistic Investigation of the Reactions between Cyclohexane Carboxaldehyde and Ureido Groups
Model
reactions have been performed to explore the reactivity of
a variety of ureido groups with cyclohexane carboxaldehyde. The reaction
mechanism between ureido and aldehyde functionalities is more complicated
than expected. A new heterocyclic product was identified, which is
very stable and solvent resistant. The final product profile is reactant
and solvent dependent. In the cases of urea, alkyl urea, and benzyl
urea, the reaction pathway goes from hemiaminal to aminal, to enamine,
and finally to the heterocyclic product with nearly 100% yield. For
other investigated ureido groups, the reaction stopped at the enamine
product, and products are a mixture of hemiaminal, aminal, and enamine.
All reaction steps are reversible except for the last step. The structure
of the unique cyclic product was determined combining NMR and LC–MS
analysis, and the reaction pathway was verified by kinetics studies
Organic Acid Quantitation by NeuCode Methylamidation
We have developed a multiplexed quantitative
analysis method for
carboxylic acids by liquid chromatography high resolution mass spectrometry.
The method employs neutron encoded (NeuCode) methylamine labels (<sup>13</sup>C or <sup>15</sup>N enriched) that are affixed to carboxylic
acid functional groups to enable duplex quantitation via mass defect
measurement. This work presents the first application of NeuCode quantitation
to small molecules. We have applied this technique to detect adulteration
of olive oil by quantitative analysis of fatty acid methyl amide derivatives,
and the quantitative accuracy of the NeuCode analysis was validated
by GC/MS. Currently, the method enables duplex quantitation and is
expandable to at least 6-plex analysis
Segmentation of Precursor Mass Range Using “Tiling” Approach Increases Peptide Identifications for MS<sup>1</sup>‑Based Label-Free Quantification
Label-free quantification is a powerful tool for the
measurement
of protein abundances by mass spectrometric methods. To maximize quantifiable
identifications, MS<sup>1</sup>-based methods must balance the collection
of survey scans and fragmentation spectra while maintaining reproducible
extracted ion chromatograms (XIC). Here we present a method which
increases the depth of proteome coverage over replicate data-dependent
experiments without the requirement of additional instrument time
or sample prefractionation. Sampling depth is increased by restricting
precursor selection to a fraction of the full MS<sup>1</sup> mass
range for each replicate; collectively, the <i>m</i>/<i>z</i> segments of all replicates encompass the full MS<sup>1</sup> range. Although selection windows are narrowed, full MS<sup>1</sup> spectra are obtained throughout the method, enabling the collection
of full mass range MS<sup>1</sup> chromatograms such that label-free
quantitation can be performed for any peptide in any experiment. We
term this approach “binning” or “tiling”
depending on the type of <i>m</i>/<i>z</i> window
utilized. By combining the data obtained from each segment, we find
that this approach increases the number of quantifiable yeast peptides
and proteins by 31% and 52%, respectively, when compared to normal
data-dependent experiments performed in replicate
Metabolism of Multiple Aromatic Compounds in Corn Stover Hydrolysate by <i>Rhodopseudomonas palustris</i>
Lignocellulosic
biomass hydrolysates hold great potential as a
feedstock for microbial biofuel production, due to their high concentration
of fermentable sugars. Present at lower concentrations are a suite
of aromatic compounds that can inhibit fermentation by biofuel-producing
microbes. We have developed a microbial-mediated strategy for removing
these aromatic compounds, using the purple nonsulfur bacterium Rhodopseudomonas palustris. When grown photoheterotrophically
in an anaerobic environment, R. palustris removes most of the aromatics from ammonia fiber expansion (AFEX)
treated corn stover hydrolysate (ACSH), while leaving the sugars mostly
intact. We show that R. palustris can
metabolize a host of aromatic substrates in ACSH that have either
been previously described as unable to support growth, such as methoxylated
aromatics, and those that have not yet been tested, such as aromatic
amides. Removing the aromatics from ACSH with R. palustris, allowed growth of a second microbe that could not grow in the untreated
ACSH. By using defined mutants, we show that most of these aromatic
compounds are metabolized by the benzoyl-CoA pathway. We also show
that loss of enzymes in the benzoyl-CoA pathway prevents total degradation
of the aromatics in the hydrolysate, and instead allows for biological
transformation of this suite of aromatics into selected aromatic compounds
potentially recoverable as an additional bioproduct
Lignin Conversion to Low-Molecular-Weight Aromatics via an Aerobic Oxidation-Hydrolysis Sequence: Comparison of Different Lignin Sources
Diverse
lignin samples have been subjected to a catalytic aerobic
oxidation process, followed by formic-acid-induced hydrolytic depolymerization.
The yield of monomeric aromatic compounds varies depending on the
lignin plant source and pretreatment method. The best results are
obtained from poplar lignin isolated via a acidolysis pretreatment
method, which gives 42 wt% yield of low-molecular-weight aromatics.
Use of other pretreatment methods and/or use of maple and maize lignins
afford yields of aromatics ranging from 3 to 31 wt%. These results
establish useful references for the development of improved oxidation/depolymerization
protocols